The theory that red blood cells (RBCs) generate and release nitric oxide (NO)-like bioactivity has gained considerable interest. However, it remains unclear whether it can be produced by endothelial NO synthase (eNOS), which is present in RBCs, and whether NO can escape scavenging by hemoglobin. The aim of this study was to test the hypothesis that arginase reciprocally controls NO formation in RBCs by competition with eNOS for their common substrate arginine and that RBC-derived NO is functionally active following arginase blockade. We show that rodent and human RBCs contain functional arginase 1 and that pharmacological inhibition of arginase increases export of eNOS-derived nitrogen oxides from RBCs under basal conditions. The functional importance was tested in an ex vivo model of myocardial ischemiareperfusion injury. Inhibitors of arginase significantly improved postischemic functional recovery in rat hearts if administered in whole blood or with RBCs in plasma. By contrast, arginase inhibition did not improve postischemic recovery when administered with buffer solution or plasma alone. The protective effect of arginase inhibition was lost in the presence of a NOS inhibitor. Moreover, hearts from eNOS −/− mice were protected when the arginase inhibitor was given with blood from wildtype donors. In contrast, when hearts from wild-type mice were given blood from eNOS −/− mice, the arginase inhibitor failed to protect against ischemia-reperfusion. These results strongly support the notion that RBCs contain functional eNOS and release NO-like bioactivity. This process is under tight control by arginase 1 and is of functional importance during ischemia-reperfusion.is a biological messenger that is a key regulator of cardiovascular function by inducing vasodilation, inhibition of platelet aggregation, and leukocyte adhesion (1). Reduced bioavailability of endothelium-derived NO is closely associated with development of several cardiovascular diseases including atherosclerosis, ischemia-reperfusion injury, and hypertension. The vascular effects of NO have traditionally been considered to be mediated by endothelium-derived NO after formation by the constitutively expressed endothelial NO synthase (eNOS). An alternative source of NO is nitrite that can be converted to NO in cardiac tissue during ischemia or hypoxia (2-4). In 1996, Stamler and colleagues suggested a role for red blood cells (RBCs) in exporting NO bioactivity and regulating blood flow (5). In this model, RBCs contain NO in the form of S-nitrosylated hemoglobin, which is in equilibrium with small nitrosothiols that are exported preferentially under deoxygenated conditions (5, 6). RBCs thereby provide NO-based vasodilatory activity through S-nitrosothiols when deoxygenated. It was also suggested that the source of RBC NO is eNOS (5). However, it was assumed that eNOS was exclusively vascular in origin, and mechanisms regulating RBC formation and export of NO bioactivity have been a matter of significant debate over the years (7). Another mechanism for ...

Inhibition of arginase protects from myocardial infarction by a mechanism that is dependent on NOS activity and bioavailability of NO by shifting arginine utilization from arginase towards NOS. These findings suggest that targeting of arginase is a promising future therapeutic strategy for protection against myocardial IR injury.

Myocardial ischaemia–reperfusion injury can be significantly reduced by an episode(s) of ischaemia–reperfusion applied prior to or during myocardial ischaemia (MI) to peripheral tissue located at a distance from the heart; this phenomenon is called remote ischaemic conditioning (RIc). Here, we compared the efficacy of RIc in protecting the heart when the RIc stimulus is applied prior to, during and at different time points after MI. A rat model of myocardial ischaemia–reperfusion injury involved 30 min of left coronary artery occlusion followed by 120 min of reperfusion. Remote ischaemic conditioning was induced by 15 min occlusion of femoral arteries and conferred a similar degree of cardioprotection when applied 25 min prior to MI, 10 or 25 min after the onset of MI, or starting 10 min after the onset of reperfusion. These RIc stimuli reduced infarct size by 54, 56, 56 and 48% (all P < 0.001), respectively. Remote ischaemic conditioning applied 30 min into the reperfusion period was ineffective. Activation of sensory nerves by application of capsaicin was effective in establishing cardioprotection only when elicited prior to MI. Vagotomy or denervation of the peripheral ischaemic tissue both completely abolished cardioprotection induced by RIc applied prior to MI. Cardioprotection conferred by delayed remote postconditioning was not affected by either vagotomy or peripheral denervation. These results indicate that RIc confers potent cardioprotection even if applied with a significant delay after the onset of myocardial reperfusion. Cardioprotection by remote preconditioning is critically dependent on afferent innervation of the remote organ and intact parasympathetic activity, while delayed remote postconditioning appears to rely on a different signalling pathway(s).

Bulhak AA, Jung C, Ö stenson C, Lundberg JO, Sjö quist P, Pernow J. PPAR-␣ activation protects the type 2 diabetic myocardium against ischemia-reperfusion injury: involvement of the PI3-kinase/Akt and NO pathway. Am J Physiol Heart Circ Physiol 296: H719 -H727, 2009. First published January 16, 2009 doi:10.1152/ajpheart.00394.2008.-Several clinical studies have shown the beneficial cardiovascular effects of fibrates in patients with diabetes and insulin resistance. The ligands of peroxisome proliferatoractivated receptor-␣ (PPAR-␣) reduce ischemia-reperfusion injury in nondiabetic animals. We hypothesized that the activation of PPAR-␣ would exert cardioprotection in type 2 diabetic Goto-Kakizaki (GK) rats, involving mechanisms related to nitric oxide (NO) production via the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. GK rats and agematched Wistar rats (n Ն 7) were given either 1) the PPAR-␣ agonist WY-14643 (WY), 2) dimethyl sulfoxide (DMSO), 3) WY and the NO synthase inhibitor N G -nitro-L-arginine (L-NNA), 4) L-NNA, 5) WY and the PI3K inhibitor wortmannin, or 6) wortmannin alone intravenously before a 35-min period of coronary artery occlusion followed by 2 h of reperfusion. Infarct size (IS), expression of endothelial NO synthase (eNOS), inducible NO synthase, and Akt as well as nitrite/ nitrate were determined. The IS was 75 Ϯ 3% and 72 Ϯ 4% of the area at risk in the Wistar and GK DMSO groups, respectively. WY reduced IS to 56 Ϯ 3% in Wistar (P Ͻ 0.05) and to 46 Ϯ 5% in GK rats (P Ͻ 0.001). The addition of either L-NNA or wortmannin reversed the cardioprotective effect of WY in both Wistar (IS, 70 Ϯ 5% and 65 Ϯ 5%, respectively) and GK (IS, 66 Ϯ 4% and 64 Ϯ 4%, P Ͻ 0.05, respectively) rats. The expression of eNOS and eNOS Ser1177 in the ischemic myocardium from both strains was increased after WY. The expression of Akt, Akt Ser473, and Akt Thr308 was also increased in the ischemic myocardium from GK rats following WY. Myocardial nitrite/nitrate levels were reduced in GK rats (P Ͻ 0.05). The results suggest that PPAR-␣ activation protects the type 2 diabetic rat myocardium against ischemia-reperfusion injury via the activation of the PI3K/Akt and NO pathway.peroxisome proliferator-activated receptor-␣ ligand; endothelial function; phosphatidylinositol 3-kinase

Reperfusion through thrombolysis or percutananeous coronary angioplasty is standard treatment in impending acute myocardial infarction. Although restoration of blood flow to the jeopardised myocardial area is a perquisite for myocardial salvage, reperfusion itself may lead to accelerated and additional myocardial injury beyond that generated by ischemia alone. This is referred to as the "reperfusion injury". Since the reperfusion injury is initiated by the treatment of myocardial infarction, it is of importance to limit the extent of the injury. Several studies aimed at preventing reperfusion injury by means of pharmacological agents have therefore been conducted. The design of such studies is crucial for the results. Factors of importance are the timing of drug administration, animal species used, the degree of collateral flow and the duration of ischemia. A variety of pharmacological compounds have been investigated in different experimental models of myocardial ischemia and reperfusion. These include oxygen free radical scavengers, antioxidants, calcium channel blockers, inhibitors of neutrophils, nitric oxide, adenosine-related agents, inhibitors of the renin-angiotensin system, endothelin receptor antagonists, Na(+)/H(+) exchange inhibitors, and anti-apoptotic agents. All these groups of pharmacological agents have been demonstrated to protect from reperfusion injury determined as limitation of infarct size, improved myocardial and endothelial function, and reduced incidence of arrhythmias. The mechanism behind the protective effect may differ between different groups of compounds, but some compounds may exert cardioprotection via common pathways. Such a pathway may be via maintained bioavailability of nitric oxide.

Abstract-The functional inhibitory characteristics of the angiotensin II type 1 receptor blockers (ARB) candesartan; irbesartan; and losartan and its active metabolite EXP 3174 (EXP) were studied in rabbit aortic strips and rat portal vein preparations in vitro. Moreover, plasma-protein binding was determined, and the binding was high (Ͼ98.5%) for all ARBs. These values were needed to relate the concentrations of the ARBs used in vitro to the nonprotein bound concentrations in clinical use. In both vascular preparations, candesartan caused a marked decrease in the maximal contractile response of the angiotensin II (Ang II) concentration-response curve. Losartan, EXP, and irbesartan caused a rightward parallel shift without any major effects on the maximal response to Ang II. The inhibitory effect of candesartan developed slowly (maximal effect after Ͼ30 minutes) and lasted Ͼ2 hours despite repeated washing of the vessels. The effect of losartan, irbesartan, and EXP had a faster onset, and most of the inhibitory effect disappeared after washing. The duration of the inhibitory effects of the ARBs were not related to lipophilicity of the compounds. Cooling of the rat portal vein preparations to 4°C before administration of candesartan prevented the persistent inhibition of Ang II response seen at 37°C. For the other ARBs studied, the magnitude of inhibition and the speed of recovery of the Ang II response were independent of the incubation temperature before washing. In addition, when candesartan was given to conscious rats, the inhibitory effect on Ang II-induced blood pressure responses persisted during the 24-hour period despite nondetectable plasma concentrations of candesartan at 24 hours. [1][2][3] The tolerability of these compounds seems to be better than that of other antihypertensives, whereas the blood pressure-lowering effect is about the same as for angiotensin-converting enzyme inhibitors and calcium antagonists. 1-3 Regarding efficacy of different ARBs, candesartan and irbesartan were shown to be more effective than losartan in lowering 24-hour blood pressure (BP) in mildly to moderately hypertensive patients. 4,5 Candesartan was described to dissociate slowly from AT 1 -receptors in cell membrane preparations and to cause a more persistent inhibition of the angiotensin II (Ang II)-mediated vascular contractile response when compared with losartan. 6 In isolated vascular preparations from the rabbit, losartan caused a parallel rightward shift in the concentration-effect curves for Ang II, 7 whereas candesartan caused a marked suppression of the maximum contractile response to Ang II. 1,6 Thus ARBs may differ in their antagonism of the Ang II-mediated response, causing surmountable or insurmountable antagonism. 8 The mechanism of the insurmountable antagonism of candesartan and the long-lasting duration of effect is not clear, although it may be related to its slow dissociation from the receptor. 1,6 The aim of our study was to investigate the functional inhibitory characteristics of different ARBs. ...

Clevidipine is an ultrashort-acting vasoselective calcium antagonist under development for short-term intravenous control of blood pressure. Studies in animals, healthy volunteers and patients have demonstrated the vascular selectivity and rapid onset and offset of antihypertensive action of clevidipine, a synthetic 1,4-dihydropyridine that inhibits L-type calcium channels. Clevidipine has a high clearance (0.05 L/min/kg). and is rapidly hydrolyzed to inactive metabolites by esterases in arterial blood. Its half-life in patients undergoing cardiac surgery is less than one min.Unlike sodium nitroprusside, a drug commonly used for the short-term control of blood pressure, which dilates both arterioles and veins, clevidipine reduces blood pressure through a selective effect on arterioles. As documented in animals and in cardiac surgical patients, clevidipine reduces peripheral resistance without any undesirable effect on cardiac filling pressure. It increases stroke volume and cardiac output. In anesthetized patients undergoing cardiac surgery clevidipine, unlike sodium nitroprusside, does not increase heart rate.In addition of having a favorable hemodynamic profile, suitable for rapid control of blood pressure, clevidipine protects against ischemia/reperfusion injuries, which are not uncommon during major surgery. In anesthetized pigs, clevidipine reduced infarct size after 45 min-long myocardial ischemia by 40%. In rats, renal function and splanchnic blood flow were better maintained when blood pressure was reduced with clevidipine than with sodium nitroprusside.

AimsAlthough the nature of the humoral factor which mediates cardioprotection established by remote ischaemic conditioning (RIc) remains unknown, parasympathetic (vagal) mechanisms appear to play a critical role. As the production and release of many gut hormones is modulated by the vagus nerve, here we tested the hypothesis that RIc cardioprotection is mediated by the actions of glucagon-like peptide-1 (GLP-1).Methods and resultsA rat model of myocardial infarction (coronary artery occlusion followed by reperfusion) was used. Remote ischaemic pre- (RIPre) or perconditioning (RIPer) was induced by 15 min occlusion of femoral arteries applied prior to or during the myocardial ischaemia. The degree of RIPre and RIPer cardioprotection was determined in conditions of cervical or subdiaphragmatic vagotomy, or following blockade of GLP-1 receptors (GLP-1R) using specific antagonist Exendin(9–39). Phosphorylation of PI3K/AKT and STAT3 was assessed. RIPre and RIPer reduced infarct size by ∼50%. In conditions of bilateral cervical or subdiaphragmatic vagotomy RIPer failed to establish cardioprotection. GLP-1R blockade abolished cardioprotection induced by either RIPre or RIPer. Exendin(9–39) also prevented RIPre-induced AKT phosphorylation. Cardioprotection induced by GLP-1R agonist Exendin-4 was preserved following cervical vagotomy, but was abolished in conditions of M3 muscarinic receptor blockade.ConclusionsThese data strongly suggest that GLP-1 functions as a humoral factor of remote ischaemic conditioning cardioprotection. This phenomenon requires intact vagal innervation of the visceral organs and recruitment of GLP-1R-mediated signalling. Cardioprotection induced by GLP-1R activation is mediated by a mechanism involving M3 muscarinic receptors.